RAP-RO

rAPid Alkaline Phosphatase

1 units/μg protein, optimum reaction temp. 37 °C

• Synonym(s): alkaline phosphatase, cloning of dna fragments
• Enzyme Commission number: 3.1.3.1 (BRENDA, IUBMB)
• UNSPSC Code: 12352204
• NACRES: NA.21

Properties

• biological source: bovine intestine
• Quality Level: 100
• recombinant: expressed in Pichia pastoris
• form: liquid
• specific activity: 1 units/μg protein
• mol wt: monomer 56 kDa by SDS-PAGE
• packaging: pkg of 1,000 U (04898133001 [1 U/μl]); pkg of 5,000 U (04898141001 [1 U/μl])
• manufacturer/tradename: Roche
• concentration: <0.1 % (w/w)
• parameter: 37 °C optimum reaction temp.
• technique(s): molecular cloning: suitable
• color: colorless
• pH: 7.6-8.5 (68 °F)
• solubility: water: miscible
• suitability: suitable for molecular biology
• NCBI accession no.: QBZ39068
• UniProt accession no.: P09487; ; P09487
• application(s): life science and biopharma
• foreign activity: Nicking activity, none detected (up to 20 µl enzyme using pBR322-DNA/ 16h / 37°C); unspecific endonuclease, none detected (up to 20 µl enzyme using Lambda-DNA 16 h/37 °C)
• storage temp.: −20°C
• Gene Information: bovine ... ALPI(280993)

Description

General description

rAPid Alkaline Phosphatase is an alkaline phosphatase isolated from bovine intestine and supplied as a recombinant enzyme of 56 kD, expressed in the yeast, Pichia Pastoris. The recombinant form ensures consistency and safety. rAPid Alkaline Phosphatase catalyzes the dephosphorylation of 5′ phosphates from DNA and RNA, nucleotides, and proteins. Alkaline phosphatase treatment prevents self-ligation of DNA fragments by removing the 5′-phosphoryl termini required by ligases; this feature is of major importance in cloning strategies to decrease vector background. Unlike calf intestinal phosphatase, rAPid Alkaline Phosphatase is rapidly, completely, and irreversibly inactivated by heat treatment for two minutes at +75 °C. It is therefore, an excellent alternative to Shrimp Alkaline Phosphatase. In addition, the enzyme is active in restriction enzyme buffers. Therefore, restriction enzyme digestion, dephosphorylation, enzyme inactivation, ligation, or 5′-end labeling can be performed without purification steps.

Specificity

Alkaline Phosphatase catalyzes the hydrolysis of numerous phosphate esters, such as esters of primary and secondary alcohols, saccharides, cyclic alcohols, phenols and amines. Phosphodiesters do not react. The enzyme hydrolyzes inorganic pyrophosphate. The kinetic properties of the enzyme depend on many factors, such as purity of enzyme, concentration of enzyme in the assay, buffer, pH etc.
Heat inactivation: rAPid Alkaline Phosphatase is inactivated by incubation at 75 °C for two minutes.

Application

• Use rAPid Alkaline Phosphatase in numerous applications, including: Removal of 5′-phosphoryl groups from nucleic acids
• Preparation of templates for 5′-end labeling
• Clean-up of PCR products by removal of dNTPs
• Dephosphorylation of proteins

Note: rAPid Alkaline Phosphatase is very rapidly inactivated by heating at +75 °C for two minutes. No additional purification steps are required after restriction and dephosphorylation. The dephosphorylated DNA can directly be used in a ligation reaction.
Note: rAPid Alkaline Phosphatase is also included in the Rapid DNA Dephos & Ligation Kit, which enables fast and efficient dephosphorylation and ligation of sticky- or blunt-end DNA fragments. With this convenient kit, dephosphorylation of blunt and sticky 5′-overhanging ends is performed in ten minutes at +37 °C and ligation is completed in just five minutes at +15–25 °C.

Features and Benefits

• Eliminate contamination risk: Recombinant expression, and reliable and sensitive quality control tests ensure high purity.
• Replace your current phosphatases with an economical solution: rAPid Alkaline Phosphatase performs equally as well as Shrimp Alkaline Phosphatase or Bovine Alkaline Phosphatase.
• Eliminate the risk of animal-derived diseases: Expression in Pichia pastoris ensures consistency and safety.
• Benefit from rapid heat inactivation: rAPid Alkaline Phosphatase is inactivated after only 2 minutes at +75°C.
• Obtain consistent performance: rAPid Alkaline Phosphatase has excellent stability during storage compared to other alkaline phosphatases.

Packaging

1 kit containing 2 components

Quality

All lots of rAPid Alkaline Phosphatase are tested for the absence of deoxyribonucleases and nicking activity according to the current Quality Control procedures.

Unit Definition

One unit of rAPid Alkaline Phosphatase is the enzyme activity which hydrolyzes 1 mol of 4-nitrophenyl phosphatase in one minute at +37 °C under assay conditions.

Volume Activity: 1 U/μl

Other Notes

For life science research only. Not for use in diagnostic procedures.

Kit Components Only

• rAPid Alkaline Phosphatase Buffer 10x concentrated
• rAPid Alkaline Phosphatase 1 U/μl

Safety Information

• Storage Class Code: 12 - Non Combustible Liquids
• WGK: WGK 1
• Flash Point(F): does not flash
• Flash Point(C): does not flash